In the last project period, this lab discovered a novel slow effect of olivocochlear (OC) stimulation that may be responsible for OC- mediated protection from acoustic trauma. The slow effect of OC stimulation is distinguished from the fast effect by a delay (10 sec), a much longer buildup of suppression, a much longer time constant of decay (30 sec), and by a diminution in magnitude with continued OC activation. Both are mediated by activation of the medial OC fibers to outer hair cells, through a neuronal-nicotinic cholinergic receptor containing the alpha-9 subunit. Patterns of OC stimulation which produce the slow effect also protect against temporary threshold shifts from acoustic over-stimulation, thus they may be linked. The goal of this project is to determine the mechanism by which the slow effect provides protection from acoustic over-stimulation and, consequently, to infer molecular mechanisms underlying the response of the hair cells to acoustic over-stimulation and the intrinsic mechanisms present in the inner ear for minimizing that damage. Ultimately, understanding the mechanism by which the ear can protect itself from acoustic trauma could lead to the development of techniques for identifying those most at risk for acoustic injury or for decreasing the ear~s vulnerability to acoustic over-stimulation. The OC system may also be involved in the development of conditioning-related protective effects, whereby repeated exposure to non-traumatic sound renders the ear less vulnerable to subsequent acoustic injury. The proposed experiments probe whether conditioning involves chronic modifications in the slow-effect cascade. If so, these results will provide the groundwork on which to pursue investigations of this broad class of phenomena in a more rational way. The proposed experiments should also have significance in the understanding the basic biology of cell-signaling. The findings of the previous project period suggest that a unique cell-signaling scheme has been implemented in outer hair cells whereby a single receptor produces both rapid and long-term effects via the unusual subcellular anatomy of these cells. This findings suggests functional roles for two unusual subcellular structures of the hair cell: the synaptic cisternae at the medial OC-outer hair cell synapse and the highly elaborated network of subsurface cisternae which line the lateral membrane of the outer hair cells in all mammals.